PROJECT SUMMARY ? PROJECT 2 Glioblastoma (GBM) has a dismal prognosis. Based on recent favorable results with other forms of cancer, immunotherapies are increasingly being tested in clinical trials of GBM. One type of immunotherapy consists of oncolytic viruses (OVs0 that exert their effects by direct tumor cytotoxicity with rounds of intratumoral infectious spread that elicit an antitumor immune response. In 2015 the first OV, based on oncolytic Herpes Simplex Virus (oHSV), was approved by the FDA. Over the last several years, we have preclinically developed a novel oHSV with GBM selectivity, designated as rQNestin34.5. The main difference between rQNestin34.5 and previous oHSVs that have been tested in human GBM clinical trials is that the HSV1 ?2 34.5 gene that encodes for the ICP34.5 protein is retained under transcriptional control of a nestin promoter/enhancer element highly expressed in GBM and in the GBM ?stem-like? cell population, but not expressed in the adult human brain. Extensive preclinical efficacy, toxicology and biodistribution studies have been carried out with rQNestin34.5 in mice, culminating in an IND (#16380) in 2016. We are now poised to initiate a ?first-in-man' Phase I clinical trial of rQNestin34.5 for recurrent GBM, which we plan to finish by the end of this proposed project. Our overarching hypothesis is that rQNestin34.5 in humans leads to evidence of an immune response and that a combination of oHSV-intratumoral oncolysis/ immunostimulation with inhibition of immune checkpoint signaling will provide increased effectiveness in animal models of glioma. We propose to test this hypothesis with the following 3 aims: Aim 1: Test if there is a maximum tolerated dose (MTD) and pilot correlative efficacy for a ?first-in-man? clinical trial of rQNestin34.5; Aim 2: Test if immune checkpoint (IC) blockade augments oHSV antiGBM effects in mice where T cells express high levels of PD-1; and Aim 3: Test if expression of IC antibodies by newly engineered oHSVs improves efficacy/toxicity profiles when compared to systemic IC antibody administration. As part of this PPG, we also plan to collaborate with Project 4 for the correlative science required for immune cell function in human specimens and with Project 3 for the science related to Notch signaling. Our trial will inform Project 1 in its design of the next generation of oHSV clinical trials. We will employ the services of Core 3 for biostatistical analyses. We will employ the services of Core 1 to generate oHSV and Core 2 for the use of glioma models. Finally, Core A provides the overall administrative functions required to integrate and manage this project with others.